Important roles for the ATP sensitive potassium (K-ATP) channel in heart have been shown for myocardial preservation and for arrhythmogenesis in ischemia. The block of the current throughout this K+ channel by ATP is a distinguishing feature. Recent studies have indicated that K-ATP can be formed by co-expressing two proteins: an inwardly rectifying channel known as KIR6.2 and a sulfonylurea receptor (SUR). The key question as to how ATP blocks this channel has not been resolved. A long term objective of this research is to elucidate the molecular mechanisms and structures of the K-ATP subunits accounting for ATP block. The central hypothesis is that the K-ATP block mechanism involves a series of conformational changes that is initiated by ATP binding to a specific site on the sulfonylurea receptor. This conformational change is then transmitted to the inwardly rectifying channel and leads to block of the current. The goals of this proposal are to identify the ATP binding site, and to determine how the ATP binding signal is transmitted from the binding site to the blocking site. The specific aims of this research are: (1) To define the isoform specific differences in ATP block using heterologously expressed K-ATP channels, (2) to identify the minimal structural requirements for the ATP blocking activity, (3) to identify on SUR, using site-directed mutations, the sequences that are critical to ATP binding, (4) to identify on both SUR and KIR6.2, using a chimeric strategy sequence, regions which are most critical for the functional coupling and subsequently to find and map, using site-directed mutagenesis, the ATP binding signal transmission pathway and the blocking site in the ionic pore.